Studies on the protein-bound chondroitin sulphate of bovine cortical bone

A fraction containing chondroitin sulphate, isolated from bovine cortical bone under mild conditions, was separated by ion-exchange chromatography into three fractions with apparent homogeneity on electrophoresis and ultracentrifugation. Two of these appeared to consist of chondroitin sulphate bound to a glycoprotein ;core' that had similarities to the bone sialoprotein described previously. The differences in composition of the two fractions were considered to be due to variation in the number or lengths of the polysaccharide chains. The presence of xylose and the alkali-lability of the bond between protein and polysaccharide suggested the presence of a xylosylserine linkage. The third fraction had the properties of a relatively pure chondroitin sulphate which contained a small amount of peptide. These fractions differed considerably from the protein-polysaccharide complexes of epiphysial and other cartilages, and their relevance to the possible role of glycosaminoglycans is discussed.     

Some studies on the composition of bovine cortical-bone sialoprotein

1. An analysis of bovine bone sialoprotein, a homogeneous glycoprotein isolated from cortical bone, is presented. 2. Analytical results agree with earlier physical measurements indicating a molecular weight of about 23000. 3. Mild acid hydrolysis and treatment with neuraminidase showed that fucose and sialic acid occupy terminal positions on oligosaccharide chains. 4. Treatment of the sialic acid-free glycoprotein with beta-galactosidase showed that much of the galactose occupies a sub-terminal location in the intact glycoprotein. 5. The polypeptide chain is rich in aspartic acid, glutamic acid, serine, threonine and glycine, and has no detectable free terminal amino group. 6. Glycopeptides were studied after proteolytic digestion. 7. It is considered that the carbohydrate moiety is highly branched and is probably linked by an acid- and alkali-stable glycosylamine bond involving aspartic acid.     

Observations on bioluminescence in some deep-water anthozoans

The luminous responses to electrical stimulation of isolated polyps of 4 deep-water anthozoans are described. All show facilitatory responses and summation at stimulus frequencies > 2 s^–1. The responses of the gorgonian Acanella arbuscula comprise a slow summation of weak individual flashes. It is suggested that there is no fundamental difference between deep and shallow species, nor between the responses of scyphozoans, such as Pelagia and Atolla, and anthozoans, such as those described here. Both facilitated and decremental responses can be obtained from each of the 2 groups and the complexity of in vivo responses may be as much a reflection of selective pressures on the neural pathways as on the bioluminescent systems themselves.     

The carboxyl terminus of the smooth muscle myosin light chain kinase is expressed as an independent protein, telokin.

It has been proposed that the carboxyl terminus of the smooth muscle myosin light chain kinase is expressed as an independent protein. This protein has been purified from tissues and named telokin (Ito, M., Dabrowska, R., Guerriero, V., Jr., and Hartshorne, D. J. (1989) J. Biol. Chem. 264, 13971-13974). In this study we have isolated and characterized cDNA and genomic clones encoding telokin. Analysis of a genomic DNA clone suggests that the mRNA encoding telokin arises from a promoter which appears to be located within an intron of the smooth muscle myosin light chain kinase (MLCK) gene. This intron interrupts exons encoding the calmodulin binding domain of the kinase. The amino acid sequence deduced from the cDNA predicts that telokin is identical to the carboxyl-terminal 155 residues of the smooth muscle MLCK. Unlike the smooth muscle MLCK which is expressed in both smooth and non-muscle tissues, telokin is expressed in some smooth muscle tissues but has not been detected in aortic smooth muscle or in any non-muscle tissues.     

Basic residues are important for Ca2+/calmodulin binding and activation but not autoinhibition of rabbit skeletal muscle myosin light chain kinase

Several allosterically modulated protein kinases have been shown to be regulated by an autoinhibitory domain located within the kinase molecules. The inhibitory domain has been proposed to act as a "pseudosubstrate" inhibitor binding to the substrate binding site of the kinase, thereby blocking the binding of the enzyme's true substrate. In this report, site-directed mutagenesis has been used to further investigate the mechanism of activation of the inhibitory domain of rabbit skeletal muscle myosin light chain kinase. Basic residues within the pseudosubstrate domain (572-573, 577-579, 580-581), which are analogous to the important substrate determinants of the myosin light chain, were found not to be required in order to maintain the kinase in an inhibited state. Two groups of these residues (577-579 and 581-582) were, however, found to be important for high affinity calmodulin binding to the kinase. These data suggest that the autoinhibitory domain of myosin light chain kinase may not function by directly mimicking the light chain substrate.     

Rapid recovery of gonadotroph function after down-regulation of receptors for GnRH in ewes

Several characteristics of the hypothalamo-hypophysial axis were examined after down-regulation of GnRH receptors and the desensitization which accompanies it in the ewe. Down-regulation of GnRH receptors, induced by i.v. infusion of GnRH (2.5 micrograms/h) for 24 h, resulted in a 50% decrease in the number of receptors for GnRH at the end of the infusion period. The number of receptors for GnRH was restored to control values by 6 h after the infusion ended and remained stable at 12, 24, 48, 72 and 96 h after infusion. The amount of LH released in response to an i.v. injection of 100 micrograms GnRH was reduced by 82% at the end of the infusion period, but there was no significant reduction in the GnRH-induced release of FSH. The GnRH-induced release of LH was restored by 12 h after the infusion ended; however, the amount of FSH released in response to GnRH was not different from control values at any time. A decrease in both the amplitude and frequency of endogenous pulses of LH was observed from 0 to 12 h after the end of the infusion period. At no time did the concentration of gonadotrophins in the pituitary change. These results demonstrate that replenishment of receptors for GnRH and recovery of the ability of the gonadotroph to release LH are associated events. However, the GnRH-induced release of FSH does not appear to be closely related to the number of GnRH receptors. We suggest that continuous exposure to GnRH may inhibit the hypothalamic pulse generator as well as the pituitary response to the pulse generator.